Early Alzheimer's disease (AD) symptoms include mild memory loss and cognitive impairment that slowly progresses to severe dementia. Clinical symptomology may initially be due to synaptic dysfunction, followed by more profound morphological changes that may include neuritic dystrophy, synaptic loss, and frank cell death. The fundamental mechanism underlying this insidiously progressive pathophysiology is thought to be an age- related accumulation of amyloid beta-protein (Abeta) fibrils, ultimately observed as mature amyloid plaques at autopsy. However, the focus on end-stage tissue has led to the assumption that fibrils per se underlie the progression of AD. Our working hypothesis is that prefibrillar, oligomeric forms of Abeta can initiate neuronal dysfunction and can directly and/or via further transition to higher molecular weight assemblies (amyloid fibrils), trigger neuronal loss. In support of this, stable Abeta oligomers have been identified in the cerebrospinal fluid of AD patients, prefibrillar forms of Abeta cause synaptic dysfunction and neuronal death, and soluble Abeta levels in brain correlate relatively well with cognitive impairment. Furthermore, transgenic mice producing human Abeta show electrophysiological changes prior to any plaque formation. Our recently published studies have shown that protofibrils (PFs), a metastable oligomeric form of Abeta, can acutely increase the electrical activity of cortical neurons and reproducibly induce neurotoxicity. In this proposal, our 4 aims focus on; 1) ionic mechanisms of altered electrophysiological activity induced by sub-lethal (nM) protofibrillar Abeta; 2) biochemical mechanisms mediating PF-induced neurotoxicity; 3) isolation and characterization of endogenous protofibrils; and 4) long-term changes in neuronal function caused by PF-induced alterations in specific gene expression. Primary mixed brain cultures will be used to assess the PF-induced electrical activity and neuronal injury. Using whole-cell patch-clamp electrophysiology, the role of calcium and potassium channels in mediating PF-induced activity will be assessed with specific channel antagonists. Early neuronal dysfunction will be assessed using subtle morphological, biochemical, and gene-expression markers. Our hypothesis and data suggest that the preclinical and clinical progression of AD is driven, in part, by early temporal changes occurring in Abeta oligomerization, not just amyloid fibril formation. Deciphering and inhibiting the biological activity of PF is a novel approach that should help in elucidating the role of early Abeta intermediates in AD and in designing rational therapeutic strategies to slow or block the progression of neuronal injury.